Aim 1:The rationale for testing further this vector in a macaque model is several folds. 1). ALVAC is an avian vector; therefore pre-existing immunity is not a factor that could enhance HIV infectivity. This is particularly important now in the face of recent results with the adenovirus based HIV vaccine trial (STEP trial) whereby five fold increase transmission of HIV was observed in individuals (not circumcised) that had high levels of prior immunity to the vector. 2). The results of the phase III trial with an ALVAC HIV vaccine III in combination with a gp120 protein boost will become available in 2009. If any protection is scored in the 16,000 volunteers, and we demonstrate in the present study an improvement on the ALVAC-SIV based vaccine immunogenicity in combination with DNA, our data are likely to influence the design of future clinical trials. 3). We need to test the improved vector in macaques because the readout of protection in the SIVmac251 macaque model is now proposed by several investigators as a gold standard for the go ahead to further clinical trials ( Summit Vaccine meeting on March 25). 4). DNA/ALVAC-SIV vaccine has not been tested in the SIVmac251 macaque model and several preclinical studies using poxvirus vaccine candidates for HIV have shown enhanced immunogenicity and protection when macaques were primed with DNA. In the recent Vaccine Summit organized by the NIAID director, Anthony Fauci,a clear consensus emerged on the utility of using SIVSM in challenge studies to test the breadth of protection from high virus level/disease induced by T-cells. An extreme point of view (in my opinion) proposed by some scientist is the idea of using SIVSM the as a gate keeper for decision to move vaccines to human trials. To date, very few studies in nonhuman primates have addressed directly the issue of HIV genetic variability. Our previous studies have demonstrated an acceptable level of protection from disease in a rigorous viral challenge (SIVmac251) macaque model using a combination of vaccine modalities and, at this time, it seems appropriate to assess the degree of protection, if any, induced by a polyvalent DNA prime/ALVAC-SIV boost vaccine regimen. The scientific goals of this aim are to improve the immunogenicity of an ALVAC-SIV vaccine in a DNA prime /ALVAC-SIV boost and to assess cross-clade protection by testing a SIVmac251-based vaccine against a mucosal challenge with a homologous and heterologous virus, SIVSME660. The preclinical data obtained in this study will begin to address the issue of cross-clade protection and perhaps provide a proof of concept on whether protection from disease by T cell vaccines is attainable against a divergent virus. In one group of animals we will define the level of protection from homologous challenge with SIVmac251, and it is expected to be equivalent as in the previous study. In the second ,immunized in a manner identical to that of the first group we will challenge intrarectally with SIVSME660 (15;16), kindly provided by V.M. Hirsch. The end points of this study will be immunological, virological, and clinical. If a reasonable degree of protection is observed strategies to further ameliorate immunogenicity of this approach (adjuvanted DNAs, CD40L expressed by ALVAC etc) will be explored. In aim 1 we wish to test the hypothesis that continuous expression of low level of SIV antigen(s) in the gut may maintain a sufficient level of effector CD8 memory cells able to decrease early seeding of the virus and sufficient level of central memory cells that may limit the broadcasting of the virus at distal sites. To do so we will explore the potential usefulness of an HIV vaccine vectored in the HTLV-II backbone. HTLV-II, is a human retrovirus that does not cause disease neither in healthy nor in HIV-I infected individuals. We have obtained evidence that indeed HTLV-II infects macaques and replicates at very low level in lymphoid tissue and particularly in the gut. Importantly HTLV-II infects dendritic cells both in vivo and in vitro and HTLV-II infected dendritic cells have a mature phenotype. We are currently preparing constructs of HTLV-II that express either the SIV Gag or fragments of SIV envelope to test the hypothesis proposed above. Aim 2 :we plan to develop an HTLV-II vectored HIV vaccine and explore the potential of HPV as a delivery system for an HIV vaccine Most of the T-cell vaccines developed for HIV are based on microbial vectors that have limited replication capacity and do not persist in the host. HTLV-II persist in the and replicates at very low level in the gut of the host. thus an HTLV--II vectored vaccine that replicate at mucosal sites might provide constitutive and effective T-cell responses as well as induce mucosal humoral immunity. HPV infects,in the appropriate conditions, the vaginal mucosa. Because most transmission of HIV worldwide are heterosexual,The HPV vaccine platform may be ideal to induce mucosal T-cell responses